NMR-based determination of disulfides in two μ-selenoconotoxin SIIIA analogs. NMR spectra at 15 °C for 1 mM SIIIA[C3U,C13U,15N/13C enriched C4 and C19] prepared in 40 mM NaPi (pH 6.2), 50 mM NaCl, 90% H2O and 10% D2O is shown in panels a and b, and SIIIA[15N/13C enriched C3 and C13,C4U,C19U] in identical solution conditions is shown in panels c and d. Panels a and c show the 2D [13C,1H] constant time HSQC and panels b and d the 2D 13C-F2-edited [1H,1H] NOESY. The proton dimensions (abscissa) are aligned for panel pairs a/b and c/d. Non-degenerate C4 and C19 CβH2’s are connected with a line (shown in panel a), the degenerate C3 and C13 CβH2’s are shown in panel c. NOE crosspeaks confirming the C4-C19 and C3-C19 disulfides are boxed. Intraresidue NOEs can be easily traced in the figure. A few natural abundance signals are present in the NOESYs but posed no problems with interpretation.